45 2 Organoids as a model to study intestinal and liver dysfunction in severe malnutrition Figure 5. Mitochondrial dysfunction after prolonged amino acid starvation in mature hepatic organoids. Mature hepatic organoids were grown in complete culture medium throughout (control, grey), amino-acid-free medium for 48 hours (48 hours starvation, blue) and starved for 96h (96 hours starvation, pink). (A) Representative immunoblot images. (B) Protein levels relative to β-actin. Quantification of data shown in (A) Data represent mean ± SEM from 3 biological replicates (biological replicates are obtained from independent experiments) (*P<0.05, ** P< 0.01, one-way ANOVA with Tukey’s post-hoc test). (C) Oxygen consumption rate in mature hepatic organoids measured as basal respiration (State 1) (ADP), State 3 (stimulated with Palmitoyl Carnitine and Malate and ADP) and uncoupled respiration (State U). Data represents the mean of 3 biological replicates (from independent experiments) ± SEM. (*P<0.05, ** P< 0.01 paired t test) Re-introduction of amino acids for 48 hours following 48 hours of starvation recovered the PMP-70 levels to almost those of the control organoids. PGC1-α and catalase levels increased, but were not significantly different from either control or starved organoids (Figure 4 a, b). Since no regulation was observed in mitochondrial protein levels after 48 hours, re-introduction of amino acids did not have any further effect. Due to the limited lifespan of the organoids in culture, re-supplementation after 96 hours of amino acid starvation could not be done.
RkJQdWJsaXNoZXIy MTk4NDMw